Environmental Microbiology (2003) 5(8), 681–690 © 2003 Society for Applied Microbiology and Blackwell Publishing Ltd Blackwell Science, LtdOxford, UKEMIEnvironmental Microbiology1462-2920Blackwell Publishing Ltd, 20035 8681690Original ArticlePhycoerythrin biodiversity in natural Prochlorococcus populationsC. Steglich, A. F. Post and W. R. Hess Received and accepted 19 February, 2003; *For correspondence. E-mail hessw@neb.com; Tel. (+1) 978 927 5054; Fax (+1) 978 921 1527. † Present address: Ocean Genome Legacy, 32 Totes Road, Beverley, MA 01915 USA. Analysis of natural populations of Prochlorococcus spp. in the northern Red Sea using phycoerythrin gene sequences Claudia Steglich, 1 Anton F. Post 2 and Wolfgang R. Hess 1 * † 1 Humboldt-University, Department of Biology, Chausseestr. 117, D-10115 Berlin, Germany. 2 H. Steinitz Marine Biology Laboratory, Interuniversity Institute for Marine Sciences, Coral Beach, POB 469, Eilat 88103, Israel. Summary Marine cyanobacteria of the genus Prochlorococcus belong to one of two ecotypes that are specifically adapted to either low light (LL) or high light (HL) con- ditions. Previous analyses of the differences in pig- mentation and gene complement revealed that LL- adapted ecotypes carry a gene cluster to produce a functional phycoerythrin, whereas in the fully sequenced genome of the HL-adapted strain MED4, only a single and free-standing cpeB gene occurs. This gene encodes a derived form of b-phycoerythrin, the function of which has remained enigmatic so far. Here, an analysis of HL-adapted Prochlorococcus strains from different ocean provinces revealed the presence of a cpeB gene highly similar to that of MED4. To investigate whether the presence of partic- ular phycoerythrin genes is a common characteristic of the LL- and HL-adapted ecotypes, primer sets tar- geting specific motifs in LL-cpeB and HL-cpeB were designed for polymerase chain reaction (PCR) analy- sis of Red Sea phytoplankton. A major PCR product for Prochlorococcus HL-cpeB was obtained from samples taken at 5–70 m depth and for LL- cpeB from 70–125 m. The high sensitivity of this approach allowed the detection of HL-cpeB down to 100 m and LL-cpeB as deep as 175 m. DNA sequence and phy- logenetic analysis of 70 individual clones for HL- cpeB and of 68 clones for LL-cpeB revealed a monophyletic origin for the HL and LL sequences respectively. This study shows that cpeB sequences are suitable as very sensitive molecular markers for the study of natural populations of Prochlorococcus. The low sequence divergence of HL-cpeB among Prochloro- coccus strains, which have been isolated from the Mediterranean Sea, the Arabian Sea and the Southern Pacific Ocean as well as in populations from the Red Sea, suggests the HL-cpeB gene to be conserved and its product to be functional in Prochlorococcus. Introduction Prochlorococcus, one of the dominant cyanobacteria in the world’s oceans, comprises an unusual pigment com- position. In contrast to the vast majority of cyanobacteria, it possesses divinyl-chlorophyll a and b that are bound to distinct chlorophyll b-binding proteins, the major antenna proteins (Garczarek et al., 2000). These proteins form a concentric ring around photosystem I (Bibby et al., 2001) and have functionally fully replaced phycobilisomes as a light-harvesting system. Consequently, phycobiliprotein genes are in a rapid evolutionary process within the genus Prochlorococcus. All genes encoding phycocyanin, allo- phycocyanin and the respective linker proteins have been lost except phycoerythrin genes, which have been selec- tively retained (Hess et al., 1996; 1999; 2001). Thus, the analysis of the evolutionary history of Prochlorococcus’ light-harvesting proteins provides an interesting system to study the replacement of one major light-harvesting sys- tem by another, and to follow the phylogenetic diversifica- tion driven by this process. The occurrence of at least two ecotypes of Prochlorococcus that differ in their depth dis- tributions, relative chlorophyll fluorescence and adaptation to either low light (LL) or high light (HL) was demonstrated in both cultures and natural populations investigated in different parts of the world (Moore et al., 1998; Urbach et al., 1998; West and Scanlan, 1999; West et al., 2001). These HL- or LL-adapted ecotypes fall into easily distin- guishable clusters in a 16S ribosomal RNA phylogenetic tree (Urbach et al., 1998), and recent evidence has con- firmed that these two ecotypes possess a very different gene complement (Garczarek et al., 2000; Hess et al., 2001). Intriguingly, some laboratory strains possess active phycoerythrin genes, among them cpeB and cpeA encod- ing the b and a subunit of phycoerythrin, as well as several genes with similarity to those encoding other phycobilip-